Gene/Protein Disease Symptom Drug Enzyme Compound
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Humans have now been growing and storing enough food for a long enough time that some rapidly evolving organisms, such as fungi, are moving into niches created by the exploitation of certain plants as food. Food is expected to be nutritious. The most important of the physicochemical conditions that affects fungal growth is related to the biological state of the food. Living foods, particularly fresh fruits, vegetables, and also grains and nuts before harvest, possess powerful defense mechanisms against microbial invasion. When the specific microorganisms overcome defense mechanisms, the spoilage of a living food starts. Other factors to consider are water activity, hydrogen ion concentration, temperature, gas tension, consistency, nutrient status, specific solute effect, and preservation. The consequences of mold contamination of foods are diverse: unsightly appearance, chemical (removal or change of most of the constituents) and nutritional value changes, modification of organoleptic quality, difficulties in preservation, occupational hazards (mycoses, allergies), and toxicoses (mycotoxicoses). It is possible to recognize a succession of three distinct mycoflora during the storage of cereals, but they can also be mixed: 1. Field fungi growing and established before harvesting (Alternaria, Fusarium, Helminthosporium, Cladosporium). 2. Storage fungi taking over and dominanting in the silo (Aspergillus and Penicillium). 3. Advanced decay fungi (Papulospora, Sordaria, Fusarium graminearum, and members of the order Mucorales).
Methods Mol Biol 2004
PMID:Fungal isolation and enumeration in foods. 1515 24

To clarify mechanisms of rice blast resistance in rice plants we used suppression subtractive hybridization (SSH) to isolate genes induced upon rice blast inoculation in a rice blast-resistant mutant. A total of 26 rice cDNAs were isolated and found to have elevated expression upon rice blast infection in a rice blast-resistant derivative, SHM-11, of the rice cultivar, Sanghaehyanghyella. Sequencing of the cDNAs revealed that many of the proteins they encoded had been previously described as involved in plant responses against pathogen attack. Two interesting groups of the defense-related proteins consisted of three different PR5 homologues and four different protease inhibitors, all highly expressed in the rice blast mutant. Genes encoding proteins involved in signal transduction and regulation were also identified, including translation initiation factor eIF5A, C2 domain DNA binding protein, putative rice EDS and putative receptor like kinase. Most of the identified cDNAs were highly expressed 24 h after blast inoculation. Our results suggest that a pathway regulating defense gene expression may be altered in the mutant, resulting in early induction of the defense genes upon fungal infection.
Mol Cells 2004 Jun 30
PMID:Identification of rice genes induced in a rice blast-resistant mutant. 1523 21

Pisatin is an isoflavonoid phytoalexin synthesized by pea (Pisum sativum L.). Previous studies have identified two enzymes apparently involved in the synthesis of this phytoalexin, isoflavone reductase (IFR), which catalyzes an intermediate step in pisatin biosynthesis, and (+)6a-hydroxymaackiain 3-O-methyltransferase (HMM), an enzyme catalyzing the terminal step. To further evaluate the involvement of these enzymes in pisatin biosynthesis, sense- and antisense-oriented cDNAs of Ifr and Hmm fused to the 35s CaMV promoter, and Agrobacterium rhizogenes, were used to produce transgenic pea hairy root cultures. PDA, a gene encoding pisatin demethylating activity (pda) in the pea-pathogenic fungus Nectria haematococca, also was used in an attempt to reduce pisatin levels. Although hairy root tissue with either sense or antisense Ifr cDNA produced less pisatin, the greatest reduction occurred with sense or antisense Hmm cDNA. The reduced pisatin production in these lines was associated with reduced amounts of Hmm transcripts, HMM protein, and HMM enzyme activity. Hairy roots containing the PDA gene also produced less pisatin. To evaluate the role of pisatin in disease resistance, the virulence of N. haematococca on the transgenic roots that produced the lowest levels of pisatin was tested. Hairy roots expressing antisense Hmm were more susceptible than the control hairy roots to isolates of N. haematococca that are either virulent or nonvirulent on wild-type pea plants. This appears to be the first case of producing transgenic plant tissue with a reduced ability to produce a phytoalexin and demonstrating that such tissue is less resistant to fungal infection: these results support the hypothesis that phytoalexin production is a disease resistance mechanism.
Mol Plant Microbe Interact 2004 Jul
PMID:Introduction of plant and fungal genes into pea (Pisum sativum L.) hairy roots reduces their ability to produce pisatin and affects their response to a fungal pathogen. 1524 74

In Colletotrichum lagenarium, RPK1 encoding the regulatory subunit of PKA is required for pathogenicity. From the rpkl mutant that forms small colonies, we isolated three growth-suppressor mutants. All rpk1-suppressor mutants are nonpathogenic and contain amino acid changes in the PKA catalytic subunit Cpkl. To assess the roles of cyclic AMP (cAMP) signaling in detail, we generated knockout mutants of CPK1 and the adenylate cyclase gene CAC1. The cpk1 and cac1 mutants are nonpathogenic on cucumber. Interestingly, both of the mutants germinated poorly, suggesting involvement of cAMP signaling in germination. Germination defect in the cpk1 and cac1 mutants is partially rescued by incubation of the conidia at lower concentrations. Germinating conidia of the cpk1 and cac1 mutants can form appressoria, but the appressoria formed by them are nonfunctional, like those of the rpk1 mutant. Cytological analysis indicates that the appressoria of the cpk1 mutant contain larger numbers of lipid bodies compared with the wild type, whereas lipid levels in the rpk1 mutants are lower, suggesting cAMP-mediated regulation of lipid metabolism for appressorium functionality. Furthermore, the cpk1 and cacl mutants have a defect in infectious growth in plant. In C. lagenarium, Cmkl mitogen-activated protein kinase (MAPK) regulates germination, appressorium formation, and infectious growth. These results suggest that cAMP signaling controls multiple steps of fungal infection in cooperative regulation with Cmkl MAPK in C. lagenarium.
Mol Plant Microbe Interact 2004 Dec
PMID:cAMP-pKA signaling regulates multiple steps of fungal infection cooperatively with Cmk1 MAP kinase in Colletotrichum lagenarium. 1559 41

Colletotrichum acutatum infects citrus petals and induces premature fruit drop and the formation of persistent calyces. The accumulation of hormones and other growth regulators, and differential gene expression in affected flowers and young fruit, was examined following fungal infection. Ethylene evolution increased threefold and indole-3-acetic acid (IAA) accumulation was as much as 140 times. Abscisic acid (ABA) levels showed no significant response. After infection, both trans- and cis-12-oxo-phytodienoic acid increased 8- to 10-fold. No significant difference of transjasmonic acid (JA) was observed in citrus flower petals or pistils. However, a fivefold increase of cis-JA was detected. The amount of salicylic acid (SA) was elevated twofold in affected petals, but not in pistils. Northern blot analyses revealed that the genes encoding ACC oxidase or ACC synthase, and 12-oxo-phytodienoic acid (12-oxo-PDA) reductase, were highly expressed in affected flowers. The genes encoding auxin-related proteins also were upregulated. Application of 2-(4-chlorophenoxy)-2-methyl-propionic acid (clofibrate; a putative auxin inhibitor), 2,3,5-triiodobenzolic acid (an auxin transport inhibitor), or SA after inoculation significantly decreased the accumulation of the gene transcripts of auxin-responsive, GH3-like protein and 12-oxo-PDA reductase, but resulted in higher percentages of young fruit retention. The results indicate that imbalance of IAA, ethylene, and JA in C. acutatum-infected flowers may be involved in symptom development and young fruit drop.
Mol Plant Microbe Interact 2004 Dec
PMID:Induction of phytohormones and differential gene expression in citrus flowers infected by the fungus Colletotrichum acutatum. 1559 45

It has been suggested that there are at least 15 Mal d 1-related (PR10) genes in one genotype of apple (Malus x domestica Borkh). We sequenced cDNA libraries of cultivar 'Royal Gala' and identified 12 members of the Mal d 1 family, including the previously reported Mal d 1b and Mal d 1d, an allelic variant of the previously reported Mal d 1a. Eight Mal d 1 gene products were expressed in tree-ripened fruit, in either the cortex or the skin, and most of these were also expressed in leaves in response to challenge with Venturia inaequalis -a fungal disease of apple. Mal d 1 gene products were identified from a large number of different tissues. Degree of ripeness as measured by standard parameters was shown not to predict either the amount of protein able to bind to a specific monoclonal antibody 5H8, previously shown to bind to an allergenic epitope in Mal d 1b and a/d, or the amount of Mal d 1 mRNA present. Mal d 1d and Mal d 1b were the most highly expressed isoforms in 'Royal Gala', particularly in the skin of fruit, and these isoforms were also predominant in other cultivars and species of apple. Genotypes, however, differed in relative predominance of Mal d 1b and Mal d 1d. The predominantly expressed Mal d 1 genes in ripe apple fruit were translated in vivo into proteins and proteins binding to the antibody were found in all cultivars and species examined. New Mal d 1 proteins were identified that bound to the 5H8 antibody. At least two new subfamilies have been identified, and while some structural differences are predicted between groups of isoforms, the P-loop motif is identical in all except two isoforms. A role in intracellular signalling in plants is suggested and in vitro expression of the isoforms should help in assessing their relative roles in disease, allergic responses, senescence and nucleotide-, cytokinin- and brassinosteroid-binding.
Plant Mol Biol 2004 May
PMID:Characterisation of Mal d 1-related genes in Malus. 1560 87

Tissue-specific or regulated expression of transgenes is desirable in order to prevent pleiotropic side effects of putatively harmful transgene products as well as loss of energy resources due to unnecessary accumulation of transgene products. Epidermis-specific expression would be useful for many defense-related genes directed against attack by fungal pathogens that enter the plant body by direct penetration through the epidermis. In an approach to enhance resistance of wheat to the powdery mildew fungus Blumeria graminis f.sp. tritici, a novel epidermis-specific promoter was developed and used for expression of two defense-related genes. A 2.3 kb fragment of the wheat GstA1 promoter in combination with an intron-containing part of the wheat WIR1a gene was found to drive strong and constitutive transient expression in wheat epidermis. This promoter-intron combination was used for overexpression of oxalate oxidase 9f-2.8 and TaPERO peroxidase, two defense-related wheat genes expressed in inner leaf tissues. Expression studies of several transgenic lines by in situ oxalate-oxidase staining, RNA and protein blot analyses, as well as real-time PCR, demonstrated strong and constitutive transgene expression in the shoot epidermis. Transient as well as stable over-expression of the TaPERO peroxidase gene in wheat epidermis under the control of the GstA1i promoter resulted in enhanced resistance against Blumeria graminis f.sp. tritici, whereas oxalate-oxidase overexpression had no effect in either system. The data suggest that the wheat GstA1 promoter in combination with the WIR1a intron is useful for transgenic approaches to fungal disease resistance in cereals.
Plant Mol Biol 2005 Jan
PMID:Stable expression of a defense-related gene in wheat epidermis under transcriptional control of a novel promoter confers pathogen resistance. 1582 82

Animal models have been helpful in assessing a drug's potential application to treatment of humans. These controlled experiments allow description of the impact of a wide range of important treatment variables, including drug dose or concentration, dosing interval, pathogen, and host immune state. Animal models of mycoses are designed to address particular therapeutic questions. Some models are designed to screen multiple compounds in a rapid manner. Other models are selected to more closely mimic clinical infections. The following chapter will be limited to the description of a murine model of disseminated candidiasis in which in vitro results, pharmacokinetic properties, and a microbiological in vivo end point is used in the assessment of a wide variety of antifungal compounds.
Methods Mol Med 2005
PMID:Use of an animal model of disseminated candidiasis in the evaluation of antifungal therapy. 1588 38

The fungus Magnaporthe grisea, the causal agent of rice blast disease, is a major pathogen of rice and is capable of producing epidemics on other cultivated cereals, including barley (Hordeum vulgare). We explored the requirements for basal resistance of barley against a compatible M. grisea isolate using both genetic and chemical approaches. Mutants of the RAR1 gene required for the function of major resistance gene-mediated resistance and mutants of the ROR1 and ROR2 genes required for full expression of cell-wall-penetration resistance against powdery mildew pathogens were examined for macroscopic and microscopic alterations in M. grisea growth and symptoms. RAR1 contributed to resistance in epidermis and mesophyll at different stages of fungal infection dependent on the MLO/mlo-5 status. Whereas no ROR2 effect was detected, ROR1 was found to contribute to cell-wall-penetration resistance, at least in the epidermis. Application of the actin agonist cytochalasin E promoted cell wall penetration by M. grisea in a dose-dependent manner, demonstrating an involvement of the actin cytoskeleton in penetration resistance.
Mol Plant Microbe Interact 2005 May
PMID:RAR1, ROR1, and the actin cytoskeleton contribute to basal resistance to Magnaporthe grisea in barley. 1591 38

The host response to infection is the outcome of a complex interaction between a microbe and a host's innate and adaptive immune system. In this context, the role of antibody in the endemic mycoses is relatively poorly understood. Recently, a monoclonal antibody to a cell surface protein has been shown to be protective in a murine histoplasmosis model. The findings with Histoplasma capsulatum may provide a paradigm for antibody protection against endemic fungi. This paper reviews the recent data on protective antibody in histoplasmosis and previous data supporting a role for antibody in protective responses in other dimorphic fungi.
Curr Mol Med 2005 Jun
PMID:Protective antibodies and endemic dimorphic fungi. 1597 99


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